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Maxwell 16 lev rna ffpe purification kit

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The Maxwell 16 LEV RNA FFPE Purification Kit is a laboratory equipment product designed for the extraction and purification of RNA from Formalin-Fixed, Paraffin-Embedded (FFPE) tissue samples. The kit utilizes magnetic bead-based technology to isolate high-quality RNA, which can be used for various downstream applications.

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Lab products found in correlation

Ribo zero, by Illumina (3 mentions) Nextseq 500 device, by Illumina (3 mentions) Proteinase k solution, by Promega (2 mentions) Superscript vilo cdna synthesis kit, by Thermo Fisher Scientific (2 mentions) Maxwell 16 instrument, by Promega (2 mentions) Maxwell 16 ffpe plus lev dna kit, by Promega (2 mentions) Qubit dsdna br assay kit, by Thermo Fisher Scientific (2 mentions) Qiazol, by Qiagen (2 mentions) 2100 bioanalyzer, by Agilent Technologies (2 mentions) Exo cip rapid pcr cleanup kit, by New England Biolabs (2 mentions) Maxwell 16 ffpe tissue lev dna purification kit, by Promega (2 mentions) Maxwell rsc device, by Promega (1 mentions) Maxwell 16 blood purification kit, by Promega (1 mentions) Maxwell 16 lev simplyrna, by Promega (1 mentions) Maxwell 16 tissue dna purification kit, by Promega (1 mentions) Agilent 4200 tapestation, by Agilent Technologies (1 mentions) Maxwell 16 ffpe plus lev dna purification kit, by Promega (1 mentions) Hotstar taq polymerase, by Qiagen (1 mentions) Bigdye terminator v3.1 cycle sequencing kit, by Thermo Fisher Scientific (1 mentions) 3730 dna analyzer, by Thermo Fisher Scientific (1 mentions)

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Maxwell 16 (79 citations) RNA purification kit (10 citations) Maxwell kits (2 citations)

29 protocols using maxwell 16 lev rna ffpe purification kit

1

Automated DNA and RNA Extraction

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Genomic DNA and RNA were extracted using the Promega Maxwell RSC device (Promega, Madison, USA). DNA was isolated with the Maxwell 16 Tissue DNA purification kit, the Maxwell 16 blood purification kit, or the Maxwell 16 FFPE Plus LEV DNA purification kit (Promega, Madison, USA) according to the manufacturer’s instructions. RNA was extracted with the Maxwell 16 LEV simplyRNA or the Maxwell 16 LEV RNA FFPE purification kit (Promega, Madison, USA) following the manufacturer’s protocols. Agilent 4200 TapeStation (Agilent Technologies, Santa Clara, USA) was used to determine RNA integrity numbers.
Kaulen L.D., Denisova E., Hinz F., Hai L., Friedel D., Henegariu O., Hoffmann D.C., Ito J., Kourtesakis A., Lehnert P., Doubrovinskaia S., Karschnia P., von Baumgarten L., Kessler T., Baehring J.M., Brors B., Sahm F, & Wick W. (2023). Integrated genetic analyses of immunodeficiency-associated Epstein-Barr virus- (EBV) positive primary CNS lymphomas. Acta Neuropathologica, 146(3), 499-514.
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2

RNA Extraction from Microdissected Samples

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Microdissected areas were directly incubate with 50 μl of lysis buffer PKD (Qiagen, Venlo, Netherlands) and 10 μl of proteinase K solution (Promega, Madison, WI, USA) at 56°C overnight. The day after samples were centrifuged at maximum speed for 10 min. RNA was purified using the Maxwell 16 LEV RNA FFPE Purification Kit (Promega) following manufacturer's instructions.
Civita P., Franceschi S., Aretini P., Ortenzi V., Menicagli M., Lessi F., Pasqualetti F., Naccarato A.G, & Mazzanti C.M. (2019). Laser Capture Microdissection and RNA-Seq Analysis: High Sensitivity Approaches to Explain Histopathological Heterogeneity in Human Glioblastoma FFPE Archived Tissues. Frontiers in Oncology, 9, 482.
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3

Extracting DNA and RNA from Tissues

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DNA and RNA of normal and tumor tissue were isolated using the Maxwell® 16 FFPE Tissue Plus LEV DNA Purification Kit (Promega) or the Maxwell® 16 LEV RNA FFPE Purification Kit (Promega), respectively, according to the manufacturer’s instructions. Both DNA and RNA were quantified by a fluorometric assay (Qubit, Thermo Fisher Scientific).
Hiltbrunner S., Cords L., Kasser S., Freiberger S.N., Kreutzer S., Toussaint N.C., Grob L., Opitz I., Messerli M., Zoche M., Soltermann A., Rechsteiner M., van den Broek M., Bodenmiller B, & Curioni-Fontecedro A. (2023). Acquired resistance to anti-PD1 therapy in patients with NSCLC associates with immunosuppressive T cell phenotype. Nature Communications, 14, 5154.
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4

TERT mRNA Expression Quantification in Solitary Fibrous Tumors

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Relative TERT mRNA expression was assessed by real-time quantitative reverse transcription PCR. Total RNA was isolated from the same formalin-fixed paraffin-embedded tumor samples that were used to extract DNA, by using the Maxwell® 16 LEV RNA FFPE Purification Kit (Promega, AS1260) according to the manufacturer’s protocol. To quantify TERT mRNA expression levels, 2 μg of total RNA from each sample was converted to cDNA by using the SuperScript® VILO cDNA Synthesis Kit (Invitrogen, 11754–010). Real-time quantitative reverse transcription PCR was performed in separate groups for extra-thoracic (soft tissue) and thoracic (pleural) solitary fibrous tumors. Real-time quantitative reverse transcription PCR was conducted in triplicate by using the TaqMan® Gene Expression Assays and gene-specific primers (Life Technologies) for TERT (Hs00972656_m1) and GAPDH (Hs02758991_g1), a housekeeping gene used as the endogenous standard. TERT expression levels were measured by using GAPDH expression as a reference, and relative quantification was determined by using the ΔΔCt method and log2 transformation.
Bahrami A., Lee S., Schaefer I.M., Boland J.M., Patton K.T., Pounds S, & Fletcher C.D. (2016). TERT promoter mutations and prognosis in solitary fibrous tumor. Modern pathology : an official journal of the United States and Canadian Academy of Pathology, Inc, 29(12), 1511-1522.
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5

Quantification of SOX11 mRNA Levels

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RNA was isolated using the Maxwell® 16 LEV RNA FFPE Purification Kit and the Maxwell® 16 Instrument (Promega, Madison, WI, USA). RT-qPCR was performed to quantify SOX11 mRNA levels (see Online Supplementary Material and Methods). Data were analyzed using the 2−ΔΔCp method and the mean of the SOX11 negative cases was defined as calibrator.
Federmann B., Frauenfeld L., Pertsch H., Borgmann V., Steinhilber J., Bonzheim I., Fend F, & Quintanilla-Martinez L. (2020). Highly sensitive and specific in situ hybridization assay for quantification of SOX11 mRNA in mantle cell lymphoma reveals association of TP53 mutations with negative and low SOX11 expression. Haematologica, 105(3), 754-764.
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6

Identification of PICALM–BRAF Fusion

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RT-PCR and Sanger sequencing were performed across the fusion break points to identify the exact fusion junction of PICALM–BRAF. The tissue blocks were cut into 4μm slides, and total RNA from FFPE samples was isolated using a tissue kit (Maxwell 16 LEV RNA FFPE purification kit, Promega, Madison, USA) and an automatic extractor (Maxwell MDx 16, Promega). RT-PCR was performed on GeneAmp 9700 using Hotstar Taq polymerase (Qiagen) as follows;15 minutes at 95°C for initial denaturation, then 45 cycles at 95°C for 30 seconds, 62°C for 30 seconds, and 72°C for 60 seconds, then 5 minutes at 72°C for final extension. 30 ng of cDNA synthesized from total RNA was used to analyze. All PCR products were sequenced on both strands using the same primers and BigDye Terminator v3.1 Cycle Sequencing kits and a 3730 DNA analyzer (Applied Biosystems). The specific fusion primers for PICALM–BRAF provided in S8 Table.
Yoo S.K., Lee S., Kim S.J., Jee H.G., Kim B.A., Cho H., Song Y.S., Cho S.W., Won J.K., Shin J.Y., Park D.J., Kim J.I., Lee K.E., Park Y.J, & Seo J.S. (2016). Comprehensive Analysis of the Transcriptional and Mutational Landscape of Follicular and Papillary Thyroid Cancers. PLoS Genetics, 12(8), e1006239.
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7

Macrodissection and Nucleic Acid Extraction

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A prototypical area within the center of the MCD lesion (neocortex) was identified on H&E slides and macrodissection performed by punch biopsy (pfm medical, Köln, Germany) or by hand. DNA and RNA were extracted from formalin-fixed paraffin-embedded (FFPE) tissue using the Maxwell 16 FFPE Plus LEV DNA Kit and Maxwell 16 LEV RNA FFPE Purification Kit (Promega, Madison, WI, USA), according to the manufacturer’s instructions. DNA concentration was quantified using the Qubit dsDNA BR Assay kit (Invitrogen, Carlsbad, CA, USA).
Kobow K., Jabari S., Pieper T., Kudernatsch M., Polster T., Woermann F.G., Kalbhenn T., Hamer H., Rössler K., Mühlebner A., Spliet W.G., Feucht M., Hou Y., Stichel D., Korshunov A., Sahm F., Coras R., Blümcke I, & von Deimling A. (2020). Mosaic trisomy of chromosome 1q in human brain tissue associates with unilateral polymicrogyria, very early-onset focal epilepsy, and severe developmental delay. Acta Neuropathologica, 140(6), 881-891.
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8

Transcriptome Analysis of RNA Fusions

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High-throughput analysis was performed as previously described [34 (link)]. Briefly, RNA was extracted using the Maxwell 16 LEV RNA FFPE Purification Kit (Promega Corporation). Libraries were prepared using Anchored-Multiplex-PCR with the commercially available Archer FusionPlex Sarcoma Panel (ArcherDx, Boulder, CO). The RNA input was 250 ng, and cDNA was synthesized using random primers. Libraries were quantified using qPCR (KAPA Biosystems, South San Francisco, CA, USA). Samples were sequenced on the MiSeq platform (Illumina, San Diego, CA, USA). The resulting FASTQ files were analyzed using the standard RNA-fusion workflow as implemented in the Archer Analysis Suite 5.1.3.
Bode-Lesniewska B., Fritz C., Exner G.U., Wagner U, & Fuchs B. (2019). EWSR1-NFATC2 and FUS-NFATC2 Gene Fusion-Associated Mesenchymal Tumors: Clinicopathologic Correlation and Literature Review. Sarcoma, 2019, 9386390.
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9

FFPE RNA-seq Transcriptome Analysis

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Total RNA was extracted from formalin-fixed paraffin-embedded (FFPE) tumor slices (5 × 5 µm) using the Maxwell 16 LEV RNA FFPE Purification kit (Promega) following manufacturer instructions. Libraries were prepared from 12 µl of total RNA with the TruSeq Stranded Total RNA using Ribo-Zero (Illumina) following manufacturer instructions. Once qualified, paired-end libraries were sequenced using 2 × 75 bp output on a NextSeq 500 device (Illumina).
The abundance of transcripts from RNA-seq data was quantified through the Kallisto program.24 (link) This program is based on pseudo alignment for rapidly determining the compatibility of reads with targets, without the need for alignment. The Kallisto transcript index used as reference was built from merged human cDNA and ncDNA files from GRCh37 assembly ENSEMBL. Gene-level count matrices were then created with the DESeq2 library. Low-count genes were pre-filtered by removing genes with too few reads.25 (link) Enrichr software was used to analyze Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways.26 (link),27 (link)
Fumet J.D., Richard C., Ledys F., Klopfenstein Q., Joubert P., Routy B., Truntzer C., Gagné A., Hamel M.A., Guimaraes C.F., Coudert B., Arnould L., Favier L., Lagrange A., Ladoire S., Saintigny P., Ortiz-Cuaran S., Perol M., Foucher P., Hofman P., Ilie M., Chevrier S., Boidot R., Derangere V, & Ghiringhelli F. (2018). Prognostic and predictive role of CD8 and PD-L1 determination in lung tumor tissue of patients under anti-PD-1 therapy. British Journal of Cancer, 119(8), 950-960.
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10

RNA Isolation and cDNA Synthesis from Organoids and FFPE

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RNA isolation from organoids was performed using the PicoPure Arcturus (Thermo Scientific, KIT0204) kit method. Tissue RNA was extracted using standard protocol of Qiazol (Qiagen) tissue lysis by TissueLyser (2 min, 20 Hz). Quality of RNA was assessed by Bioanalyzer (Agilent). RNA from FFPE material was extracted using the Maxwell® 16 LEV RNA FFPE Purification Kit (Promega, AS1260). Total RNA was used for cDNA synthesis using random primers and RNAse H-MML-V reverse transcriptase first-strand cDNA synthesis system (Promega AG, Dübendorf, Switzerland). For qPCR, cDNA (10 ng per reaction) was amplified in a CFX Real Time Detection system (Bio-Rad, Cressier, Switzerland) using SYBR Green Supermix reagent (Bio-Rad). Expression levels were normalized to the transcripts of HPRT and ACTB. Primer sequences are indicated in Supplementary Table 5.
Karkampouna S., La Manna F., Benjak A., Kiener M., De Menna M., Zoni E., Grosjean J., Klima I., Garofoli A., Bolis M., Vallerga A., Theurillat J.P., De Filippo M.R., Genitsch V., Keller D., Booij T.H., Stirnimann C.U., Eng K., Sboner A., Ng C.K., Piscuoglio S., Gray P.C., Spahn M., Rubin M.A., Thalmann G.N, & Kruithof-de Julio M. (2021). Patient-derived xenografts and organoids model therapy response in prostate cancer. Nature Communications, 12, 1117.
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